M. Rikkers (Utrecht, NL)
UMC Utrecht OrthopaedicsPresenter Of 2 Presentations
12.4.7 - Platelet lysate for cartilage regeneration: Improved redifferentiation after expansion, but opposite effects in 3D culture
Abstract
Purpose
The purpose of this study is to define the potential of platelet lysate as an animal-free alternative to FBS in expansion of human chondrocytes for cell therapy purposes. Additionally, the effects of using platelet lysate for redifferentiation of chondrocytes in 3D pellets are assessed.
Methods and Materials
Platelet lysate was prepared from human blood and platelet enrichment was measured. Chondrocytes were isolated from human osteoarthritic cartilage (age 55-84, mean 66 years) and subjected to a range of concentrations of platelet lysate in monolayer culture. Cell proliferation and morphology were assessed after 7 days. In addition, expression of chondrogenic genes was determined by quantitative real-time PCR. Next, platelet lysate-expanded chondrocytes were cultured in 3D cell pellets and cartilage matrix production was assessed after 28 days. Glycosaminoglycan (GAG) and collagen production in the pellets were quantified by biochemical analyses as well as (immuno)histochemistry. Besides, platelet lysate was supplemented into redifferentiation medium of chondrocytes in 3D pellets. Cartilage matrix production was assessed with similar outcomes.
Results
Platelet lysate had a dose-dependent effect on chondrocyte proliferation, but expression of chondrogenic markers was decreased in monolayers when compared to serum-expanded monolayers. GAG production after 28 days of subsequent 3D pellet culture was significantly higher in pellets consisting of chondrocytes expanded with 1% platelet lysate compared to controls (figure 1). When used for redifferentiation of chondrocyte pellets, platelet lysate significantly decreased the production of GAGs and collagen. This was confirmed by (immuno)histochemistry.
Conclusion
In conclusion, low concentrations of platelet lysate stimulate chondrocyte proliferation in 2D monolayer and cartilage ECM production in subsequent 3D pellet culture. However, this does not work for high concentrations of platelet lysate. Furthermore, this study showed unfavourable effects on redifferentiation of 3D chondrocyte cultures in platelet lysate. Thus, platelet lysate is a promising animal-free alternative to be used for chondrocyte expansion.
18.2.1 - Articular cartilage-derived progenitors from healthy and osteoarthritic human cartilage show potential for cartilage repair treatment
Abstract
Purpose
This study aims to isolate, characterize, and compare articular cartilage-derived progenitor cells (ACPCs) from healthy and osteoarthritic human cartilage, and to demonstrate the potential to use these ACPCs in addition to primary chondrocytes for one-stage procedures to treat cartilage defects.
Methods and Materials
Cells were isolated from full-thickness healthy (n=6, age 46-49, mean age 48) and osteoarthritic (n=6, age 41-82, mean age 62) human cartilage. Subsequently, ACPCs were isolated from the total cell population by clonal growth after a differential fibronectin adhesion assay. Healthy and osteoarthritic ACPCs were characterized by multilineage differentiation and flow cytometry analysis to assess surface marker expression. Next, ACPCs were cocultured with osteoarthritic chondrocytes in 3D pellet cultures in a range of cell-cell ratios. Pellets were harvested after 28 days and assessed for cartilage-like matrix production using quantitative biochemical analyses for glycosaminoglycans and collagen. (Immuno)histochemistry was performed to visualize proteoglycan and collagen production.
Results
Healthy and osteoarthritic ACPCs were successfully differentiated into the adipogenic and chondrogenic lineage, but failed to produce calcified matrix when exposed to osteogenic induction media. Adipogenic differentiation was more successful in osteoarthritic ACPCs, whereas chondrogenic differentiation was more distinct in healthy ACPCs. Both ACPC populations met the criteria for cell surface marker expression to identify mesenchymal stromal cells (MSCs) as shown by flow cytometry. Cartilage-like matrix production was observed in ACPC pellet cultures, as well as pellets consisting of a coculture of ACPCs and chondrocytes (figure 1).
Conclusion
In conclusion, this study provides further insight into the progenitor cell population in both healthy and osteoarthritic human articular cartilage, which shows similarities to MSCs. Furthermore, ACPCs show potential for use complementary to osteoarthritic chondrocytes in one-stage cartilage repair treatments, as a potential alternative to MSCs.